Project Summary/Abstract Periodontitis is a bacterially induced chronic inflammation of the tooth supporting tissues that may result in alveolar bone destruction and tooth loss. Tannerella forsythia (Tf), a member of the ?red-complex? bacteria group, is one of the major pathogens implicated in periodontitis. T. forsythia has been shown to co-aggregate with the ?bridge-bacterium? Fusobacterium nucleatum (Fn) to form synergistic co-biofilms in vitro and induce alveolar bone loss in mice when co- infected with Fn. The overall goal of this proposal is to determine the molecular mechanisms of T. forsythia -F. nucleatum intergeneric interactions to better understand the pathogenesis of periodontitis. Our preliminary data show that a Tf secreted -glucanase enzyme (GlcA) whose expression is induced in response to Fn sensing plays a significant role in the development of mixed biofilms. We showed that this enzyme hydrolyzes -glucans into glucose, which serves as a nutrient for Fn to promote its biomass in Tf -Fn co-biofilms. Our data showed that the increased glucose availability did not affect Tf biomass but it rather enhanced the production of methylglyoxal (MGO), a highly reactive dicabonyl compound toxic to bacterial and host cells. The goal of this proposal is to understand the mechanistic basis of Tf-Fn interactions and how metabolic interactions between these two species contribute to the development of the dental plaque, dysbiosis and inflammation. Our working hypothesis is that b-glucanase produced by Tf in response to Fn, and possible other stimuli, releases glucose from dietary b- glucans as a nutrient for the microbial community at large and as a metabolic precursor for MGO secretion to favor microbial dysbiosis. To interrogate this hypothesis, we propose two specific aims: Aim 1: To define the molecular mechanism of regulation of Tf GlcA operon in response to Fn sensing. We will analyze the mechanism and function of ECF sigma-anti sigma system that is predicted to drive Tf glcA -glucanase operon to respond stimulate with Fn and other stimuli, and; Aim 2 To determine the mechanisms by which Fn resists Tf-produced methylglyoxal and how hydrolyzed glucans impact microbial community structure. Here, we will determine the molecular mechanisms by which Fn detoxifies MGO produced by Tf in biofilms and promote dental plaque development. Successful completion of this study will form the foundation for future investigations exploring the unique ability of Tf glucanase-MGO axis in promoting microbial dysbiosis and subsequently develop glucanase targeting inhibitors to block the synergistic Tf-Fn associations and dental plaque development for the treatment of periodontitis.